Hepcidin is a recently described hormone that plays a critical role in iron metabolism. Hepcidin targets ferroportin for degradation thereby reducing export of iron from macrophages and enterocytes. Induction of hepcidin production by inflammatory mediators reduces iron availability for erythropoiesis leading to the anemia of inflammation. The PIs have combined their expertise in chemical biology and disease modeling to investigate how bone morphogenetic protein (BMP) signaling regulates hepcidin and ferroportin expression. The team's accomplishments include discovery of the first small molecule inhibitors of BMP signaling and demonstration that these inhibitors can treat anemia of inflammation in animal models. More recently, it has become apparent that the BMP-hepcidin-ferroportin signaling pathway can contribute to inflammatory diseases not typically attributed to abnormalities of iron homeostasis, including atherosclerosis. In this renewal application, the PIs propose to continue using genetics and chemical biology to study iron homeostasis, with a new focus on the role of hepcidin-ferroportin signaling in macrophage function and the role of this signaling pathway in health and disease. Three aims are proposed. In Aim 1, the PIs will take advantage of facile genetic and pharmacological manipulations in transparent zebrafish to characterize the role of hepcidin-ferroportin signaling and iron status in macrophage development, migration, and phagocytosis. In Aim 2, the PIs propose to undertake complementary approaches to elucidate the regulation of macrophage function by hepcidin-ferroportin-iron signaling in genetically-modified mice with atherosclerosis. Finally, in Aim 3, th PIs will undertake an in vivo screen using transgenic zebrafish to identify novel small molecules that stabilize ferroportin in the presence of hepcidin. Ferroportin-stabilizing compounds will be evaluated for potency, specificity, and conservation of effect in mice. The proposed research program will provide important insights into the roles of hepcidin-ferroportin-iron signaling in macrophage function and inflammatory diseases, as well as new zebrafish and mouse models and small molecule probes. Moreover, the identification of small molecules that stabilize ferroportin may represent novel therapeutic agents for the treatment of inflammatory diseases, including atherosclerotic cardiovascular disease.